Role of von Willebrand Factor in Platelet Thrombus Formation

描述（由申请人提供）：本申请中提出的研究继续致力于阐明血管性血友病因子（VWF）A1结构域（VWFA 1）中的VWF可差异调节特异性粘附功能。实验证据表明，糖蛋白（GP）Ib 1与固定在表面上的VWFA 1的相互作用，导致血小板粘附，或在溶液中，介导血小板聚集，可能通过不同的机制发生。我们的目标是确定这些不同的VWF活动的启动和调节的独特的结构决定因素，并建立其在血管损伤模型中的功能相关性。在目标2中，我们打算确定1-凝血酶是否是VWF粘附特性的生理调节剂。我们假设VWFA 1和1-凝血酶在与相同的GPIb 1受体结合时建立分子间接触，并且这独立于剪切应力影响VWFA 1-GPIb 1键的稳定性。我们建议确定的VWFA 1残基，支持与GPIb 1结合的1-凝血酶的相互作用，从而定义一个新的机制，在血栓形成过程中的VWF功能的调节。在目标3中，我们建议评估在人群中发现的特异性IgG是否是VWF活性的调节剂。我们已经确定了在人类和小鼠血液中的特异性IgG，选择性结合VWFA 1，并获得了初步证据，这种IgG可能在血栓形成中发挥作用。我们的目标是表征特异性IgG的结构，定义与VWFA 1的相互作用模式，并获得其生理病理学意义的体内证据。在目标4中，我们打算确定胶原结合的VWF导致血小板活化的信号传导功能。我们的特点是独特的胞浆内Ca++信号，血小板粘附到固定的VWFA 1或胶原蛋白在流动条件下，并发现它们增强血小板与胶原蛋白结合的VWF相互作用时。我们建议解剖的胶原蛋白-VWF复合物支持的血小板活化的机制和流体动力学的影响的过程中。这项研究的结果将提高我们的能力，影响疾病的进程，涉及血小板在动脉粥样硬化血栓形成。 公共卫生相关性： 实验和临床证据表明VWF在血栓形成中的关键作用，通过与其他蛋白质的相互作用进行协调，并通过流动血液产生的力进行调节。了解这些不同的功能需要解决结构细节以及验证的概念，在体内模型。这将导致更好的诊断和治疗心血管和心血管疾病。

DESCRIPTION (provided by applicant): The studies proposed in this application continue efforts to elucidate von Willebrand factor (VWF) in the VWF A1 domain (VWFA1) can differentially regulate specific adhesive functions. Experimental evidence indicates that the interaction of glycoprotein (GP) Ib1 with VWFA1 immobilized onto a surface, leading to platelet adhesion, or in solution, mediating platelet aggregation, may occur through different mechanisms. Our goal is to define the distinctive structural determinants underlying the initiation and regulation of these diverse VWF activities and establish their functional relevance in models of vascular injury. In aim 2 we intend to ascertain whether 1-thrombin is a physiologic modulator of VWF adhesive properties. We hypothesize that VWFA1 and 1-thrombin establish inter- molecular contacts when bound to the same GPIb1 receptor, and this influences the stability of the VWFA1-GPIb1 bond independently of shear stress. We propose to identify the VWFA1 residues that support the interaction with GPIb1-bound 1-thrombin, thus defining a novel mechanism for the regulation of VWF function during thrombogenesis. In aim 3 we propose to evaluate whether a specific IgG found in the human population is a modulator of VWF activity. We have identified in human and mouse blood a specific IgG that binds selectively to VWFA1, and obtained preliminary evidence that this IgG may play a role in thrombus formation. Our goal is to characterize the structure of the specific IgG, define the mode of interaction with VWFA1 and obtain definitive in vivo evidence for its physiopathological significance. In aim 4 we intend to define the signaling function of collagen- bound VWF leading to platelet activation. We have characterized the distinctive intracytoplasmic Ca++ signals that follow platelet adhesion to immobilized VWFA1 or collagen under flow conditions, and found that they are enhanced when platelets interact with collagen-bound VWF. We propose to dissect the mechanisms of platelet activation supported by the collagen-VWF complex and the effects of hydrodynamic force on the process. The results of this research will improve our ability to influence disease processes that involve platelets in atherothrombosis. PUBLIC HEALTH RELEVANCE: Experimental and clinical evidence points to a key role of VWF in thrombus formation, orchestrated by interactions with other proteins and modulated through forces generated by flowing blood. Understanding these different functions requires addressing structural details as well as verification of concepts in intravidal models. This will lead to a better diagnosis and treatment for cardio- and cerebro-vascular diseases.